Liposomes and Lipid Droplets Display a Reversal of Charge-Induced Hydration Asymmetry.

The unique properties of water are critical for life. Water molecules have been reported to hydrate cations and anions asymmetrically in bulk water, being a key element in the balance of biochemical interactions. We show here that this behavior extends to charged lipid nanoscale interfaces. Charge hydration asymmetry was investigated by using nonlinear light scattering methods on lipid nanodroplets and liposomes. Nanodroplets covered with negatively charged lipids induce strong water ordering, while droplets covered with positively charged lipids induce negligible water ordering. Surprisingly, this charge-induced hydration asymmetry is reversed around liposomes. This opposite behavior in charge hydration asymmetry is caused by a delicate balance of electrostatic and hydrogen-bonding interactions. These findings highlight the importance of not only the charge state but also the specific distribution of neutral and charged lipids in cellular membranes.

Charge transfer across C-H⋠⋠⋠O hydrogen bonds stabilizes oil droplets in water.

The hydrophobic­water interface plays a key role in biological interactions. However, both the hydrophobic­water interfacial molecular structure and the origin of the negative zeta potential of hydrophobic droplets in water are heavily contested. We report polarimetric vibrational sum-frequency scattering of the O­D and C-H stretch modes of 200-nanometer hexadecane oil droplets dispersed in water. An unusually broad spectral distribution (2550 to 2750 per centimeter) of interfacial water molecules that were not hydrogen bonded to other water molecules was observed, as well as a blue shift in the vibrational frequency of the interfacial hexadecane C-H stretch modes. Oil and water frequency shifts correlated with the negative electrostatic charge. Molecular dynamics simulations demonstrated that the unexpected strong charge-transfer interactions arose from interfacial C­H∙∙∙O hydrogen bonds.

Zn2+ Binds to Phosphatidylserine and Induces Membrane Blebbing.

Herein, we show that Zn2+ binds to phosphatidylserine (PS) lipids in supported lipid bilayers (SLBs), forming a PS-Zn2+ complex with an equilibrium dissociation constant of ∼100 µM. Significantly, Zn2+ binding to SLBs containing more than 10 mol % PS induces extensive reordering of the bilayer. This reordering is manifest through bright spots of high fluorescence intensity that can be observed when the bilayer contains a dye-labeled lipid. Measurements using atomic force microscopy (AFM) reveal that these spots represent three-dimensional unilamellar blebs. Bleb formation is ion specific, inducible by exposing the bilayer to µM concentrations of Zn2+ but not Mg2+, Cu2+, Co2+, or Mn2+. Moreover, Ca2+ can induce some blebbing at mM concentrations but not nearly as effectively as Zn2+. The interactions of divalent metal cations with PS lipids were further investigated by a combination of vibrational sum frequency spectroscopy (VSFS) and surface pressure-area isotherm measurements. VSFS revealed that Zn2+ and Ca2+ were bound to the phosphate and carboxylate moieties on PS via contact ion pairing, dehydrating the lipid headgroup, whereas Mg2+ and Cu2+ were bound without perturbing the hydration of these functional groups. Additionally, Zn2+ was found to dramatically reduce the area per lipid in lipid monolayers, while Mg2+ and Cu2+ did not. Ca2+ could also reduce the area per lipid but only when significantly higher surface pressures were applied. These measurements suggest that Zn2+ caused lipid blebbing by decreasing the area per lipid on the side of the bilayer to which the salt was exposed. Such findings have implications for blebbing, fusion, oxidation, and related properties of PS-rich membranes in biological systems where Zn2+ concentrations are asymmetrically distributed.

Introduction of Positive Charges into Zwitterionic Phospholipid Monolayers Disrupts Water Structure Whereas Negative Charges Enhances It.

The interfacial water structure and phosphate group hydration of 1,2-dioleoyl- sn-glycero-3-phosphatidylcholine monolayers were investigated at air/water interfaces. Both vibrational sum frequency spectroscopy (VSFS) and Langmuir monolayer compression measurements were made. The PC lipids oriented water molecules predominantly through their phosphate-choline (P-N) dipoles and carbonyl moieties. Upon the introduction of low concentrations of 1,2-dioleoyl-3-trimethylammonium propane (DOTAP), a positively charged double chain surfactant, the TAP headgroups were attracted to the phosphate moieties on adjacent PC lipids. This attraction caused the monolayers to contract, expelling water molecules that were hydrogen bonded to the phosphate groups. Moreover, amplitude of the OH stretch signal decreased. At higher DOTAP concentrations, the positive charge on the monolayer caused an increase in the area per headgroup and water molecules in the near-surface bulk region became increasingly aligned. Under these latter conditions, the OH stretch amplitude was linearly proportional to the surface potential. By contrast, introducing 1,2-dioleoyl- sn-glycero-3-phosphatidylglycerol, a negatively charged lipid, did not change the area per lipid or the phosphate-water hydrogen bonding network. As the interfacial potential grew more negative, the OH stretch amplitude increased continuously. Significantly, changes in the interfacial water spectrum were independent of the chemistry employed to create the positive or negative interfacial potential. For example, Ca2+ and tetracaine (both positively charged) disrupted the water structure similarly to low DOTAP concentrations, whereas SCN- and ibuprofen (both negatively charged) enhanced the water structure. These results suggest a direct correlation amongst the interfacial water structure, area per lipid, and surface charge density.

Multistep Interactions between Ibuprofen and Lipid Membranes.

Ibuprofen (IBU) interacts with phosphatidylcholine membranes in three distinct steps as a function of concentration. In a first step (<10 µM), IBU electrostatically adsorbs to the lipid headgroups and gradually decreases the interfacial potential. This first step helps to facilitate the second step (10-300 µM), in which hydrophobic insertion of the drug occurs. The second step disrupts the packing of the lipid acyl chains and expands the area per lipid. In a final step, above 300 µM IBU, the lipid membrane begins to solubilize, resulting in a detergent-like effect. The results described herein were obtained by a combination of fluorescence binding assays, vibrational sum frequency spectroscopy, and Langmuir monolayer compression experiments. By introducing trimethylammonium-propane, phosphatidylglycerol, and phosphatidylethanolamine lipids as well as cholesterol, we demonstrated that both the chemistry of the lipid headgroups and the packing of lipid acyl chains can substantially influence the interactions between IBU and the membranes. Moreover, different membrane chemistries can alter particular steps in the binding interaction.

Plexcitons: The Role of Oscillator Strengths and Spectral Widths in Determining Strong Coupling.

Strong coupling interactions between plasmon and exciton-based excitations have been proposed to be useful in the design of optoelectronic systems. However, the role of various optical parameters dictating the plasmon-exciton (plexciton) interactions is less understood. Herein, we propose an inequality for achieving strong coupling between plasmons and excitons through appropriate variation of their oscillator strengths and spectral widths. These aspects are found to be consistent with experiments on two sets of free-standing plexcitonic systems obtained by (i) linking fluorescein isothiocyanate on Ag nanoparticles of varying sizes through silane coupling and (ii) electrostatic binding of cyanine dyes on polystyrenesulfonate-coated Au nanorods of varying aspect ratios. Being covalently linked on Ag nanoparticles, fluorescein isothiocyanate remains in monomeric state, and its high oscillator strength and narrow spectral width enable us to approach the strong coupling limit. In contrast, in the presence of polystyrenesulfonate, monomeric forms of cyanine dyes exist in equilibrium with their aggregates: Coupling is not observed for monomers and H-aggregates whose optical parameters are unfavorable. The large aggregation number, narrow spectral width, and extremely high oscillator strength of J-aggregates of cyanines permit effective delocalization of excitons along the linear assembly of chromophores, which in turn leads to efficient coupling with the plasmons. Further, the results obtained from experiments and theoretical models are jointly employed to describe the plexcitonic states, estimate the coupling strengths, and rationalize the dispersion curves. The experimental results and the theoretical analysis presented here portray a way forward to the rational design of plexcitonic systems attaining the strong coupling limits.

What Is the Preferred Conformation of Phosphatidylserine-Copper(II) Complexes? A Combined Theoretical and Experimental Investigation.

Phosphatidylserine (PS) has previously been found to bind Cu2+ in a ratio of 1 Cu2+ ion per 2 PS lipids to form a complex with an apparent dissociation constant that can be as low as picomolar. While the affinity of Cu2+ for lipid membranes containing PS lipids has been well characterized, the structural details of the Cu-PS2 complex have not yet been reported. Coordinating to one amine and one carboxylate moiety on two separate PS lipids, the Cu-PS2 complex is unique among ion-lipid complexes in its ability to adopt both cis and trans conformations. Herein, we determine which stereoisomer of the Cu-PS2 complex is favored in lipid bilayers using density functional theory calculations and electron paramagnetic resonance experiments. It was determined that a conformation in which the nitrogen centers are cis to each other is the preferred binding geometry. This is in contrast to the complex formed when two glycine molecules bind to Cu2+ in bulk solution, where the cis and trans isomers exist in equilibrium, indicating that the lipid environment has a significant steric effect on the Cu2+ binding conformation. These findings are relevant for understanding lipid oxidation caused by Cu2+ binding to lipid membrane surfaces and will help us understand how ion binding to lipid membranes can affect their physical properties.

The complex nature of calcium cation interactions with phospholipid bilayers.

Understanding interactions of calcium with lipid membranes at the molecular level is of great importance in light of their involvement in calcium signaling, association of proteins with cellular membranes, and membrane fusion. We quantify these interactions in detail by employing a combination of spectroscopic methods with atomistic molecular dynamics simulations. Namely, time-resolved fluorescent spectroscopy of lipid vesicles and vibrational sum frequency spectroscopy of lipid monolayers are used to characterize local binding sites of calcium in zwitterionic and anionic model lipid assemblies, while dynamic light scattering and zeta potential measurements are employed for macroscopic characterization of lipid vesicles in calcium-containing environments. To gain additional atomic-level information, the experiments are complemented by molecular simulations that utilize an accurate force field for calcium ions with scaled charges effectively accounting for electronic polarization effects. We demonstrate that lipid membranes have substantial calcium-binding capacity, with several types of binding sites present. Significantly, the binding mode depends on calcium concentration with important implications for calcium buffering, synaptic plasticity, and protein-membrane association.

Unquenchable Surface Potential Dramatically Enhances Cu(2+) Binding to Phosphatidylserine Lipids.

Herein, the apparent equilibrium dissociation constant, K(Dapp), between Cu(2+) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-l-serine (POPS), a negatively charged phospholipid, was measured as a function of PS concentrations in supported lipid bilayers (SLBs). The results indicated that K(Dapp) for Cu(2+) binding to PS-containing SLBs was enhanced by a factor of 17,000 from 110 nM to 6.4 pM as the PS density in the membrane was increased from 1.0 to 20 mol %. Although Cu(2+) bound bivalently to POPS at higher PS concentrations, this was not the dominant factor in increasing the binding affinity. Rather, the higher concentration of Cu(2+) within the double layer above the membrane was largely responsible for the tightening. Unlike the binding of other divalent metal ions such as Ca(2+) and Mg(2+) to PS, Cu(2+) binding does not alter the net negative charge on the membrane as the Cu(PS)2 complex forms. As such, the Cu(2+) concentration within the double layer region was greatly amplified relative to its concentration in bulk solution as the PS density was increased. This created a far larger enhancement to the apparent binding affinity than is observed by standard multivalent effects. These findings should help provide an understanding on the extent of Cu(2+)-PS binding in cell membranes, which may be relevant to biological processes such as amyloid-ß peptide toxicity and lipid oxidation.